Our work thus far indicates that decreasing left ventricular output (LVO) by obstructing LV inflow decreases the size of the LV chamber of midgestational fetal lambs within a few days. Decreasing LVO by partially obstructing LV outflow (banding the ascending aorta) also decreases LVO but increases LV pressure by 50 percent and LV wall thickness by more than 100 percent. The aim of the research for the coming year is to determine whether the increase in LV wall thickness in response to increased afterload in the fetal lamb is related to myocardial hypertrophy (increased cell width) or hyperplasia (increased cell number), using non-banded twin fetuses as controls. We discovered that standard histological techniques could not be used. Formalin fixed, paraffin inbedded tissues exhibited massive and uncontrolled shrinkage and distortion which would have made measurements of cell width meaningless. Standard electron microscope (EM) preparation techniques preserved cellular detail but prevented resolution of the space between adjacent cells and hence demarcation of cell boundaries by light microscopy. During the past year we developed a modified technique of tissue preparation in which the beating heart is fixed by perfusion with a slightly hypertonic glutaraldehyde solution that produces a very slight increase in the extracellular space. The tissues are imbedded in methacrylate and cut into 1 micron sections and stained with Toluidine blue. Under phase-contrast light microscopy, cell boundaries are easily discerned, yet cellular and tissue architecture remains well preserved.